Simple lateral flow immunoassay devices have been developed and commercialised for detection of analytes in fluid samples, see for example EP291194. Such devices typically comprise a porous carrier comprising a dried, mobilisable labelled binding reagent capable of binding to the analyte in question, and an immobilised binding reagent also capable of binding to the analyte provided at a detection zone downstream from the labelled binding reagent. Detection of the immobilised labelled binding reagent at the detection zone provides an indication of the presence of analyte in the sample.
Alternatively, when the analyte of interest is a hapten, the immunoassay device may employ a competition reaction wherein a labelled analyte or analyte analogue competes with analyte present in the sample for binding to an immobilised binding reagent at a detection zone. Alternatively the assay device may employ an inhibition reaction whereby an immobilised analyte or analyte analogue is provided at a detection zone, the assay device comprising a mobilisable labelled binding reagent for the analyte.
An assay device may be able to detect the presence and/or amount of more than one analyte. For example, in the case of assays detecting the presence of drugs of abuse, the device may be capable of determining a whole panel of drugs. Such lateral flow immunoassay devices are generally provided with multiple detection zones, such zones being provided on a single or multiple lateral flow carriers, within the assay device.
Determination of the result of the assay has been traditionally carried out by eye. However such devices require the result to be interpreted by the user which introduces an undesirable degree of subjectivity, particularly at low analyte levels when the intensity of the detection zone is faint.
As such, digital devices have been developed comprising an optical detection means arranged to determine the result of the assay as well as a display means to display the result of the assay. Digital assay readers for use in combination with assay test-strips for determining the concentration and/or amount of analyte in a fluid sample are known as are assay devices comprising an integral digital assay reader. An example of such a device is disclosed in EP 1484601.
Light from a light source, such as a light emitting diode (LED), is shone onto a portion of the porous carrier and either reflected or transmitted light is detected by a photodetector. Typically, the reader will have more than one LED to illuminate various zones of the carrier, and a corresponding photodetector is provided for each of the plurality of LEDs. EP1484601 discloses an optical arrangement for a lateral flow test strip digital reading device comprising a baffle arrangement allowing for the possibility of reducing the number of photodetectors in the device.
Assay technology of the foregoing type has been implemented into “self-test” pregnancy testing devices. These are, typically, devices which are used by women who suspect they may be pregnant. As such, they must be designed in such a way that they are easy to use (not requiring any medical or technical training), and are typically disposable after a single use. The device is usually a lateral flow immunoassay device, and normally the test is initiated by contacting a sampling portion of a lateral flow assay stick with a urine sample. The sampling portion of the assay stick may be immersed into a sample of urine in a container or, more typically, the user may urinate directly onto the sampling portion. The assay then runs without the woman needing to perform any further steps, and the result is indicated and read by eye or, in a digital device, is determined by an assay result reading means and displayed to the user, by means of a display such as, for example, a liquid crystal display (LCD).
Conventional pregnancy tests of this sort work by measuring human chorionic gonadotrophin (hCG) in the sample. The hCG is produced by the developing embryo and a concentration of hCG in the sample above a certain threshold will trigger a positive (i.e. “pregnant”) result.
There is a need for an improved pregnancy test, especially an improved self-test pregnancy test: many women wish to know, as soon as possible, if they are pregnant, and there is a need therefore for a very sensitive test, which can detect hCG in samples, such as urine, at very low concentration. However, this creates a problem because hCG can sometimes be present in urine, at relatively low concentration, for reasons other than pregnancy, such that a very sensitive pregnancy test may give a false positive result (i.e. the specificity of the assay is diminished).
As an illustration of this, non-pregnancy associated hCG may be present in a urine sample. More especially, hCG may be present in urine from peri-menopausal and post-menopausal women, and derives from the pituitary rather than a developing embryo. The detection of this pituitary gland-derived hCG, or other non-pregnancy associated hCG, in a pregnancy test will give rise to a false positive result for pregnancy. In many countries, women defer starting a family until later in life e.g. due to work or other commitments, and so a small but significant market for ‘home’ or self-test pregnancy test devices is constituted by older women who may fall into the peri-menopausal or post-menopausal bracket, and are therefore susceptible to false positive results if using a sensitive hCG assay. According to one study, up to 10% of the sales of OTC pregnancy tests have been to women >40 yrs old (Leavitt S A 2006, “A private little revolution: the home pregnancy test in American Culture”. Bull. Hist. Med. 2006; 80:317-45).
According to the World Health Organization, the recognized definitions of “menopause” and “perimenopause” are as follows:
Menopause (natural menopause)—is defined as the permanent cessation of menstruation resulting from the loss of ovarian follicular activity. Natural menopause is recognized to have occurred after 12 consecutive months of amenorrhea, for which there is no other obvious pathological or physiological cause. Menopause occurs with the final menstrual period (FMP) which is known with certainty only in retrospect a year or more after the event.
Perimenopause—the term perimenopause includes the period immediately prior to the menopause (when the endocrinological, biological, and clinical features of approaching menopause commence) and the first year after menopause.
Accordingly, for present purposes, peri-menopausal women are defined as being those women also are in peri-menopause according to the WHO definition above and post-menopausal women are defined as those who have undergone menopause according to the WHO definition above.
Self-test or ‘home’ pregnancy tests need to be reliable to ensure women take appropriate action on receiving their test result. A desirable reliability target is ≥99% accuracy (i.e. a combined false positive and false negative rate of 1% or less). Currently available self-test or home test devices can detect urinary hCG at a concentration of 25 mIU/ml or more with a sensitivity of ≥99%. Such devices can achieve the desired ≥99% accuracy target, but only if used on the day the subject expects her period to start (i.e. expected first day of menstrual bleeding), or later, because from that time point nearly all women who are pregnant will have a urinary hCG concentration of 25 mIU/ml or greater, and levels of non-pregnancy associated hCG never normally attain this level. However, it follows that if a woman uses a self-test device before her day of expected period, a false negative result is possible because the urinary hCG concentration has not yet reached a level detectable by the test. Accordingly, currently-available conventional self-test pregnancy test devices are not 99% accurate when used before the day of expected period. In fact the median level of hCG in urine 10 days following ovulation is about 8.4 mIU/ml, and only about 10% of samples from this day would have a hCG concentration ≥25 mIU/ml. Therefore, a very low pregnancy detection rate would be seen using a conventional self-test device with 25 mIU/ml sensitivity this early in pregnancy. At 11 days following ovulation, median level rises to 19.8 mIU/ml, so at this stage less than 50% of women would be likely to receive a positive result. Detection rates for later testing would be approximately 70% (day 12), 80% (day 13) and nearly 100% (day 14), based on a 25 mIU/ml test sensitivity. Simply using a more sensitive test is not a solution, since this would increase the risk of a false positive result, due to increased likelihood of detecting non-pregnancy associated hCG, so the test would still not be ≥99% accurate.
There exists therefore a need for a pregnancy test device, especially a self-test or home test device, which is able to detect pregnancy with ≥99% accuracy, even when used at a time point earlier in pregnancy than the day of expected period.
In a different context, in many countries, tests for serum hCG are routinely performed on nearly all women patients of child-bearing age before conducting any medical intervention which might harm a developing foetus. The problem of elevated serum hCG levels, due to “pituitary” hCG, in peri- and post-menopausal women is recognised. Snyder et al., (Clinical Chemistry 2005 51, 1830-1835) examined changes with age in serum hCG concentrations of non-pregnant women and investigated the use of serum follicle-stimulating hormone (FSH) measurements as an aid to interpreting higher than expected hCG results. They suggested that a combination of serum hCG measurements, knowledge of the age of the subject, and serum FSH measurements, could be used to reduce or avoid “false positive” pregnancy results. These workers were not, however, concerned with self-test pregnancy tests and, in particular, were not concerned with detecting pregnancy at a very early stage.